PROJECT SUMMARY The purpose of this R21 is to apply cutting edge molecular immunology techniques to determine the features of the adaptive immune response likely to protect against active tuberculosis infection. Although approximately 1/3 of the world?s population is infected with tuberculosis, the vast majority of infected individuals maintain control of latent TB infections throughout their lives. We have demonstrated in prior studies that individuals with prior treated extrapulmonary TB, and no sign of residual or new infection, have increased levels of immune activation compared to individuals with treated pulmonary TB, or treated latent infection. Furthermore, these individuals have lower levels of in vitro cytokine production in response to infected macrophages. This suggests an underlying immune defect in individuals prone to disseminated TB infection. We have established cutting edge molecular techniques that allow us to 1) single-cell sort pathogen-reactive T cells 2) directly sequence their T cell receptor (TCR) alpha and beta chains 3) determine the T cell receptor alpha and beta chains of each T cell and 4) Reconstitute the alpha beta TCR complex in a reporter cell line to allow fine mapping of epitope specificity. As a proof of concept, in this application we will apply these techniques to an existing repository of peripheral blood mononuclear cells from individuals with latent tuberculosis (treated and untreated) and individuals with prior treated pulmonary or extrapulmonary TB. We hypothesize that the degree of TB-specific TCR diversity may be related to the degree of control an individual has over latent infection.